Liquid mycorrhiza compositions

ABSTRACT

The present invention relates to liquid mycorrhiza compositions and to methods for colonizing a plant, grass, tree or shrub with one or more mycorrhizas.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. application Ser. No.12/178,864 filed Jul. 24, 2008 (now allowed) which claims priority orthe benefit under 35 U.S.C. 119 of U.S. provisional application No.60/951,503 filed Jul. 24, 2007, the contents of which are fullyincorporated herein by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention relates to liquid mycorrhiza compositions and tomethods for colonizing a plant, grass, tree or shrub with one or moremycorrhizas. Specifically, it relates to compositions which improve theability of the mycorrhizas to colonize plant roots, resulting in greaterefficacy of plant treatment formulations which contain the mycorrhizas.

2. Description of Related Art

Mycorrhizas are symbiotic associations between fungi and the roots ofplants. In a mycorrhizal association, the fungus may colonize the rootsof a host plant either intercellularly, intracellularly orextracellularly. The functional symbiosis provides a suitable andsufficient carbohydrate source for the fungal symbiont. The plantsymbiont benefits can be numerous and include improved nutrient andwater uptake, additional carbon acquisition, increased sink strength forphotosynthate translocation, increased production of phytohormones,improved resistance to pathogens, and heavy metal tolerance. Mycorrhizasare critically important organs for resource uptake by most terrestrialplants. In the absence of an appropriate fungal symbiont, manyterrestrial plants suffer from resource limitations and ultimatelyreduced growth, and fitness.

Mycorrhizas are added to fertilizer products to incorporate beneficialmycorrhizas in the fertilizer blends. Current fertilizer blends containmycorrhizas in the form of a dry granular product or a wettable powderproduct. Mycorrhizas have not been added to liquid fertilizers due tostability issues.

Production of mycorrhizas is challenging because they have unique growthrequirements. Endomycorrhizas (also called vesicular arbuscularmycorrhizas, VAM, or arbuscular mycorrhizas, AM) are obligate symbiontswith plants. The predominant method used to cultivate theendomycorrhizas is with a host plant under controlled conditions.

Bactericides are commonly used to eliminate unwanted bacterialcontamination. The preservatives used at low rates are designed to killvegetative bacteria while leaving spores unharmed. Kathon(methylchloroisothiazolinone and methylisothiazolinone) and Bronopol(2-bromo-2-nitro-1,3-propanediol) are two bactericides used aspreservatives. Kathon has a broad antimicrobial spectrum. It iseffective against gram-negative and gram-positive bacteria, fungi, andyeasts. Bronopol is effective in killing gram-negative and gram-positivebacteria. Depending on the amount used, the bactericide can killbacteria or keep them in the spore form.

CN 1420167 discloses liquid inoculants basically connected withectomycorrhizas.

JP 4141023 discloses a liquid medium comprising a stabilizer (citricacid or its salt form) to protect the spores.

US 2004/0208852 discloses a method of colonizing a plant withmycorrhiza.

WO 2006/060968 discloses a liquid mycorrhizal inoculant.

Schreiner et al. (Biol. Fertil. Soils, 1997, 24:18-26) describe theeffects of biocides on arbuscular mycorrhizas.

Al-Garni (World Journal of Agricultrual Sciences, 2006, 2(3): 303-310)describes the influence of malathion and mancozeb on mycorrhizalcolonization.

West et al. (Journal of Ecology, 1993, 81(2): 345-350) describe theinfluence of three biocides on the fungal associates of the roots ofVulpia ciliate.

It is an object of the present invention to provide a liquid mycorrhizacomposition comprising mycorrhiza which increases mycorrhizalcolonization.

SUMMARY OF THE INVENTION

The present invention relates to liquid mycorrhiza compositions and tomethods for increasing mycorrhizal colonization of a plant.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to liquid compositions comprising one ormore mycorrhizas and a bactericide and/or bacteriostat, and to methodsfor increasing mycorrhizal colonization of a plant. Improvedcolonization of roots by mycorrhizas leads to better plant growth andyield.

The mycorrhiza is present in the liquid compositions in an amount of0.2-5%, preferably 0.33-3.3% weight/volume.

The bactericide and/or bacteriostat is present in an amount of 50-3,000ppm of the active ingredient, preferably 150-2,000 ppm.

Mycorrhizas

The mycorrhizas contained in the liquid compositions of the presentinvention can be any mycorrhiza. The two most common types ofmycorrhizas are ectomycorrhizas and endomycorrhizas (more commonly knownas arbuscular mycorrhizas).

Endomycorrhizas

Arbuscular mycorrhizas, or AM (formerly known as vesicular-arbuscularmycorrhizas), are an example of a mycorrhiza that involves entry of thehyphae into the plant root cell walls to produce structures that areeither balloon-like (vesicles) or dichotomously-branching invaginations(arbuscules). The fungal hyphae do not in fact penetrate the protoplast(i.e., the interior of the cell), but invaginate the cell membrane. Thestructure of the arbuscules greatly increases the contact surface areabetween the hypha and the cell cytoplasm to facilitate the transfer ofnutrients between them.

Arbuscular mycorrhizas are formed only by fungi in the divisionGlomeromycota, which are typically associated with the roots ofherbaceous plants, but may also be associated with woody plants. Fossilevidence and DNA sequence analysis suggest that this mutualism appeared400-460 million years ago, when the first plants were colonizing land.Arbuscular mycorrhizas were likely to have been very helpful at thattime, protecting plants from adverse conditions such as lack of waterand nutrients.

Arbuscular mycorrhizal fungi are quite extraordinary organisms. Firstthey have been asexual for many million years and secondly, individualscan contain many genetically different nuclei (a phenomenon calledheterokaryosis).

This type of association is found in 85% of all plant families in thewild, including many crop species such as grains.

Preferably, the endomycorrhiza is of the phylum Glomeromycota and generaGlomus and Gigaspora. In a preferred embodiment, the endomycorrhiza is astrain of Glomus, such as Glomus aggregatum, Glomus brasilianum, Glomusclarum, Glomus deserticola, Glomus etunicatum, Glomus fasciculatum,Glomus intraradices, Glomus monosporum, or Glomus mosseae, or a strainof Gigaspora margarita.

Ectomycorrhizas

Ectomycorrhizas, or EcM, typically form between the roots of woodyplants and fungi belonging to the divisions Basidiomycota, Ascomycota,or Zygomycota.

These are external mycorrhizas that form a cover on root surfaces andbetween the root's cortical cells.

Besides the mantle formed by the mycorrhizas, most of the biomass of thefungus is found branching into the soil, with some extending to theapoplast, stopping short of the endodermis.

Found in 10% of plant families, mostly woody species, including the oak,pine, eucalyptus, dipterocarp, and olive families.

Preferably, the ectomycorrhiza is of the phylum Basidiomycota. In apreferred embodiment, the ectomycorrhiza is a strain of Laccariabicolor, Laccaria laccata, Pisolithus tinctorius, Rhizopogon amylopogon,Rhizopogon fulvigleba, Rhizopogon luteolus, Rhizopogon villosuli,Scleroderma cepa, or Scleroderma citrinum.

Other Forms of Mycorrhizas

Arbuscular and ectomycorrhizas form ericoid mycorrhiza with many plantsbelonging to the order Ericales, while some Ericales form arbutoid andmonotropoid mycorrhizas. All orchids are mycoheterotrophic at some stageduring their lifecycle and form orchid mycorrhizas with a range ofbasidiomycete fungi.

The mycorrhiza may be an ericoid mycorrhiza, preferably of the phylumAscomycota, such as Hymenoscyphous ericae or Oidiodendron sp.

The mycorrhiza also may be an arbutoid mycorrhiza, preferably of thephylum Basidiomycota.

In addition, the mycorrhiza may be a monotripoid mycorrhiza, preferablyof the phylum Basidiomycota.

The mycorrhiza also may also be an orchid mycorrhiza, preferably of thegenus Rhizoctonia.

The active component of the mycorrhiza may be the spores, hyphae,extramatrix arbuscular mycelium, glomalin and rootlets, colonized by thefungus in question.

Biocides

The liquid compositions of the present invention comprise a bactericideand/or a bacteriostat. A bactericide is an agent that kills bacteria anda bacteriostat is an agent, usually chemical, that prevents the growthof bacteria but that does not necessarily kill them or their spores.

Bactericides

The bactericide for use in the liquid compositions of the presentinvention may be a disinfectant, antiseptic or antibiotic.

A bactericidal disinfectant may be:

active chlorine (i.e., hypochlorites, chloramines, dichloroisocyanurateand trichloroisocyanurate, wet chlorine, chlorine dioxide, etc.),

active oxygen (peroxides, such as peracetic acid, potassium persulfate,sodium perborate, sodium percarbonate and urea perhydrate),

iodine (iodpovidone (povidone-iodine, Betadine), Lugol's solution,iodine tincture, iodinated nonionic surfactants),

concentrated alcohols (mainly ethanol, 1-propanol, called alson-propanol and 2-propanol, called isopropanol and mixtures thereof;further, 2-phenoxyethanol and 1- and 2-phenoxypropanols),

phenolic substances (such as phenol (also called “carbolic acid”),cresols (called “Lysole” in combination with liquid potassium soaps),halogenated (chlorinated, brominated) phenols, such as hexachlorophene,triclosan, trichlorophenol, tribromophenol, pentachlorophenol, Dibromoland salts thereof),

cationic surfactants, such as some quaternary ammonium cations (such asbenzalkonium chloride, cetyl trimethylammonium bromide or chloride,didecyldimethylammonium chloride, cetylpyridinium chloride, benzethoniumchloride) and others, non-quarternary compounds, such as chlorhexidine,glucoprotamine, octenidine dihydrochloride, etc.),

strong oxidizers, such as ozone and permanganate solutions;

heavy metals and their salts, such as colloidal silver, silver nitrate,mercury chloride, phenylmercury salts, copper sulfate, copperoxide-chloride, etc. Heavy metals and their salts are the most toxic,and environment-hazardous bactericides and therefore, their use isstrongly oppressed or eliminated; further, also

properly concentrated strong acids (phosphoric, nitric, sulfuric,amidosulfuric, toluenesulfonic acids) and

alkalis (sodium, potassium, calcium hydroxides), such as of pH<1 or >13,particularly under elevated temperature (above 60° C.), kills bacteria.

A bactericidal antiseptic may be:

properly diluted chlorine preparations (e.g., Daquin's solution, 0.5%sodium or potassium hypochlorite solution, pH-adjusted to pH 7-8, or0.5-1% solution of sodium benzenesulfochloramide (chloramine B)),

some iodine preparations, such as iodopovidone in various galenics(ointment, solutions, wound plasters), in the past also Lugol'ssolution,

peroxides as urea perhydrate solutions and pH-buffered 0.1-0.25%peracetic acid solutions,

alcohols with or without antiseptic additives, used mainly for skinantisepsis,

weak organic acids such as sorbic acid, benzoic acid, lactic acid andsalicylic acid,

some phenolic compounds, such as hexachlorophene, triclosan andDibromol, and

cation-active compounds, such as 0.05-0.5% benzalkonium, 0.5-4%chlorhexidine, 0.1-2% octenidine solutions.

A bactericidal antibiotic may be penicillin, cephalosporins, andaminoglycosidic antibiotics.

Other bactericidal antibiotics include the fluoroquinolones,nitrofurans, vancomycin, monobactams, co-trimoxazole, and metronidazole.

Preferred bactericides are:

-   Halogen containing compounds such as:

Bronopol—active 2-bromo-2-nitro-1,3-propanadiol

Dowicil 75—active 1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantanechloride

DBNPA—active dibromonitrilopropionamide

-   OrganoSulfurs—includes Isothaizolones such as:

Proxel (Nipacide)—active 1,2-benzisothiazolin-3-one

Kathon—active 5-chloro-2-methyl-4-isosthiazolin-3-one,2-methyl-4-isosthiazolin-3-one

-   Nitrogen containing compounds such as:

Germall II (Diazolidinyl urea)

Tris nitro(tris(hydroxymethyl)nitromethane)

-   Phenolics such as:

Dowicide (sodium o-phenylphenate)

-   Inorganics such as:

copper arsenates

cuprous oxide

-   Organometallics such as:

compounds of arsenic, copper, mercury

-   Quaternary ammonium compounds    Bacteriostats

A bacteriostat is a biological or chemical agent that causesbacteriostasis. It stops bacteria from reproducing, while notnecessarily harming them otherwise. Upon removal of the bacteriostat,the bacteria usually start to grow again.

Bacteriostats for use in the liquid compositions of the presentinvention include sodium azide and thimerosol.

Other bactericides and bacteriostats which can be used in the liquidcompositions of the present invention are Glycopon, Ottasept (acombination of Proxel and propylene glycol), and EDTA (ethylenediaminetetraacetic acid, a chelating agent).

Other Components in the Liquid Compositions

The liquid compositions of the present invention may include one or moreof the following components:

-   1. Fertilizer ingredients:

a. Diammonium Phosphate

b. Potassium Nitrate

c. Urea

d. Potassium Phosphate

e. Potassium Bicarbonate

f. Lidoquest Fe13P

g. Lidoquest Mn13P

h. Lidoquest Zn13P

i. Potassium Chloride (KCl)

j. Disodium Dihydro Molybdate

k. Cobalt Chloride Hexahydrate

l. Nickel Chloride Hexahydrate

m. Caustic Potash Liquid

n. MKP

o. P-paraben/Nipasol

p. Potassium thiosulfate

q. Sodium hydroxide

-   2. Non-Nutritiional Plant Beneficial Substances or Biostimulants

a. Seaweed Extracts (Ascophyllum nodosum)

b. Humic Acids

c. Ascorbic Acid

d. Thiamine mononitrate

e. Potassium Sorbate (50% K Sorb)

f. Myo-Inositol

g. Glycine

h. Vitamin E (alpha tocopherol)

Methods of Colonizing a Plant with Mycorrhiza

The present invention also relates to methods of increasing thecolonization with one or more mycorrhizas comprising applying a liquidcomposition of the present invention to a plant, grass, tree or shrub.

The liquid composition can be applied in water by means of irrigationand fertigation systems.

The invention is further defined in the following paragraphs:

-   Paragraph 1. A liquid composition, comprising

(a) one or more mycorrhizas; and

(b) a bactericide and/or a bacteriostat.

-   Paragraph 2. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are ectomycorrhizas.-   Paragraph 3. The liquid composition of paragraph 2, wherein the    ectomycorrhizas are of the phylum Basidiomycota.-   Paragraph 4. The liquid composition of paragraph 3, wherein the    ectomycorrhizas are strains of Laccaria bicolor, Laccaria laccata,    Pisolithus tinctorius, Rhizopogon amylopogon, Rhizopogon fulvigleba,    Rhizopogon luteolus, Rhizopogon villosuli, Scleroderma cepa, or    Scleroderma citrinum.-   Paragraph 5. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are endomycorrhizas.-   Paragraph 6. The liquid composition of paragraph 5, wherein the    endomycorrhiza is of the phylum Zygomycota and order Glomales.-   Paragraph 7. The liquid composition of paragraph 6, wherein the    endomycorrhiza is a strain of Gigaspora margarita, Glomus    aggregatum, Glomus brasilianum, Glomus clarum, Glomus deserticola,    Glomus etunicatum, Glomus fasciculatum, Glomus intraradices, Glomus    monosporum, or Glomus mosseae.-   Paragraph 8. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are ericoid mycorrhizas.-   Paragraph 9. The liquid composition of paragraph 8, wherein the    ericoid mycorrhizas are of the phylum Ascomycota.-   Paragraph 10. The liquid composition of paragraph 9, wherein the    ericoid mycorrhizas are Hymenoscyphous ericae or Oidiodendron sp.-   Paragraph 11. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are arbutoid mycorrhizas.-   Paragraph 12. The liquid composition of paragraph 11, wherein the    arbutoid mycorrhizas are preferably of the phylum Basidiomycota.-   Paragraph 13. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are monotripoid mycorrhizas.-   Paragraph 14. The liquid composition of paragraph 13, wherein the    monotripoid mycorrhiza are of the phylum Basidiomycota.-   Paragraph 15. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are orchid mycorrhizas.-   Paragraph 16. The liquid composition of paragraph 15, wherein the    orchid mycorrhizas are of the genus Rhizoctonia.-   Paragraph 17. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are a mixture of ectomycorrhizas and    endomycorrhizas.-   Paragraph 18. The liquid composition of paragraph 1, which comprises    a bactericide.-   Paragraph 19. The liquid composition of paragraph 18, wherein the    bactericide is a disinfectant, antiseptic or antibiotic.-   Paragraph 20. The liquid composition of paragraph 19, wherein the    disinfectant is selected from the group consisting of

a. active chlorine,

b. active oxygen,

c. iodine,

d. concentrated alcohols,

e. phenolic substances,

f. cationic surfactants,

g. strong oxidizers,

h. heavy metals and their salts,

i. properly concentrated strong acids, and

j. alkalis.

-   Paragraph 21. The liquid composition of paragraph 19, wherein the    antiseptic is selected from the group consisting of

a. properly diluted chlorine preparations,

b. iodine preparations,

c. peroxides,

d. alcohols with or without antiseptic additives,

e. weak organic acids,

f. phenolic compounds, and

g. cation-active compounds.

-   Paragraph 22. The liquid composition of paragraph 19, wherein the    antibiotic is selected from the group consisting of

a. penicillin,

b. cephalosporins,

c. aminoglycosidic antibiotics,

d. fluoroquinolones,

e. nitrofurans,

f. vancomycin,

g. monobactams,

h. co-trimoxazole, and

i. metronidazole.

-   Paragraph 23. The liquid composition of paragraph 18, wherein the    bactericide is selected from the group consisting of

a. 2-bromo-2-nitro-1,3-propanadiol,

b. 1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantane chloride,

c. dibromonitrilopropionamide,

d. 1,2-benzisothiazolin-3-one,

e. 5-chloro-2-methyl-4-isosthiazolin-3-one,2-methyl-4-isosthiazolin-3-one,

f. diazolidinyl urea,

g. tris(hydroxymethyl)nitromethane,

h. sodium o-phenylphenate,

i. copper arsenates,

j. cuprous oxide,

k. compounds of arsenic, copper, mercury, and

l. quarternary ammonium compounds.

-   Paragraph 24. The liquid composition of paragraph 1, which comprises    a bacteriostat.-   Paragraph 25. The liquid composition of paragraph 24, wherein the    bateriostat is sodium azide or thimerosol.-   Paragraph 26. The liquid composition of paragraph 1, wherein the    bactericide and/or the bacteriostat is Glycopon, Ottasept or EDTA.-   Paragraph 27. The liquid composition of paragraph 1, which comprises    a bactericide and a bateriostat.-   Paragraph 28. The liquid composition of paragraph 1, wherein the one    or more mycorrhizas are present in an amount of 0.2-5%, preferably    0.33-3.3%.-   Paragraph 29. The liquid composition of paragraph 1, wherein the    bactericide and/or the bacteriostat is present in an amount of    50-3,000 ppm, preferably 150-2,000 ppm of the active indgredient.-   Paragraph 30. The liquid composition of paragraph 1, which further    comprises a fertilizer ingredient, preferably selected from the    group consisting of

a. Diammonium Phosphate,

b. Potassium Nitrate,

c. Urea,

d. Potassium Phosphate,

e. Potassium Bicarbonate,

f. Lidoquest Fe13P,

g. Lidoquest Mn13P,

h. LidoquestZn13P,

i. Potassium Chloride (KCl),

j. Disodium Dihydro Molybdate,

k. Cobalt Chloride Hexahydrate,

l. Nickel Chloride Hexahydrate,

m. Caustic Potash Liquid,

n. MKP,

o. P-paraben/Nipasol,

p. Potassium thiosulfate, and

q. Sodium hydroxide.

-   Paragraph 31. The liquid composition of paragraph 1, which further    comprises a non-nutritional plant beneficial substance or    biostimulant, preferably selected from the group consisting of

a. Seaweed Extracts (Ascophyllum nodosum)

b. Humic Acids,

c. Ascorbic Acid,

d. Thiamine mononitrate,

e. Potassium Sorbate (50% K Sorb),

f. Myo-Inositol,

g. Glycine, and

h. Vitamin E (alpha tocopherol).

-   Paragraph 32. A method of increasing the colonization with one or    more mycorrhizas, comprising applying a liquid composition of any of    paragraphs 1-31 to a plant, grass, tree or shrub.

EXAMPLES

An endomycorrhizal mean infection percentage (MIP) bioassay wasinitiated to test mycorrhizal infectivity of a liquid mycorrhizalformulation treated with bactericides. The MIP bioassay is used todetermine the effect of endomycorrhizal colonization in the roots ofcorn plants on the treatment applied. The MIP bioassay was adapted toutilize 15 mL of liquid per pot.

The liquid mycorrhizal formulation which was tested was MycoApply Ultra(Novozymes Biologicals), which is a mycorrhizal inoculant that containsseveral species of endomycorrhizas. For a ready to use fertilizer, theoptimal amount of mycorrhizas to be added based on application rate ofready to use fertilizer, mycorrhizal infectivity, and cost effectivenesswas 0.1 propagules of mycorrhizas per 1.0 mL of ready to use fertilizer.MycoApply Ultra contains 286 propagules per gram, therefore 0.00034 gMycoApply Ultra in 1 mL of ready to use fertilizer will provide thedesired concentration of mycorrhizal propagules, 0.1 prop/mL.

The MycoApply Ultra was tested with two different concentrates 10×(0.0034 g/mL) and 100× (0.034 g/mL). The bactericide used in alltreatments for Trial 1 was 500 ppm Kathon and 250 ppm Bronopol. Therewere seven treatment groups, each contained 5 replicates. Treatment 1was a water only control. Treatment 2 was 10× concentrate MycoApplyUltra treated with bactericide made 10 days before treatment. Treatment3 was 10× concentrate MycoApply Ultra treated with bactericide made 24hours before treatment. Treatment 4 was 10× concentrate MycoApply Ultrauntreated and made directly before treatment. Treatment 5 was 100×concentrate MycoApply Ultra treated with bactericide made 10 days beforetreatment. Treatment 6 was 100× concentrate MycoApply Ultra treated withbactericide made 24 hours before treatment. Treatment 7 was 100×concentrate MycoApply Ultra untreated and made directly beforetreatment. Each treatment was mixed in 100 mL of tap water.

Pots were filled within 2 cm of the top with a mix media 90:10 v/v blendof sand to peat. Two corn seeds (Zea Maize ‘Golden Bantam’) were plantedin each pot and treatment groups were randomly assigned. On the day ofplanting 15 mL of each corresponding treatment was applied to each pot.Three additional 15 mL treatments were applied weekly following theinitial treatment. The pots were placed in a growth chamber receiving 10hours of light per day (Table 1) and were watered as needed. Upongermination of the seeds the plants were thinned to one seed per pot.The corn was harvested after 4 weeks of growth.

TABLE 1 10 hour growth chamber cycle used to grow corn. a.  6:00, 20°C., 55% RH, 2 Fluorescent, 2 Incandescent b.  7:00, 22° C., 55% RH, 4Fluorescent, 2 Incandescent c. 10:00, 26° C., 65% RH, 6 Fluorescent, 8Incandescent d. 15:00, 24° C., 65% RH, 4 Fluorescent, 4 Incandescent e.18:00, 20° C., 55% RH, 0 Fluorescent, 0 Incandescent

At harvest the parameter evaluated was percent mycorrhizal colonization.

Trial 2 was set up exactly like Trial 1, but with the addition of 2 moretreatment groups. MycoApply Ultra was again tested with two differentconcentrates 10× (0.0034 g/mL) and 100× (0.034 g/mL). Trial 2 used thebactericides 500 ppm Kathon and 250 ppm Bronopol, but also added a 500ppm Kathon alone treatment. There were nine treatment groups, eachcontained 5 replicates. Treatment 1 was a water only control. Treatment2 was 10× concentrate MycoApply Ultra treated with 500 ppm Kathon and250 ppm Bronopol made 10 days before treatment. Treatment 3 was 10×concentrate MycoApply Ultra treated with 500 ppm Kathon alone made 10days before treatment. Treatment 4 was 10× concentrate MycoApply Ultratreated with 500 ppm Kathon and 250 ppm Bronopol made 24 hours beforetreatment. Treatment 5 was 10× concentrate MycoApply Ultra untreated andmade directly before treatment. Treatment 6 was 100× concentrateMycoApply Ultra treated with 500 ppm Kathon and 250 ppm Bronopol made 10days before treatment. Treatment 7 was 100× concentrate MycoApply Ultratreated with 500 ppm Kathon alone made 10 days before treatment.Treatment 8 was 100× concentrate MycoApply Ultra treated with 500 ppmKathon and 250 ppm Bronopol made 24 hours before treatment. Treatment 9was 100× concentrate MycoApply Ultra untreated and made directly beforetreatment. Each treatment was mixed in 100 mL of tap water. The rest ofthe methods follow Trial 1 exactly.

Results:

In both trials over the course of the studies plants in the 10×concentrate treatment received 0.204 g of MycoApply Ultra or 58.34mycorrhizal propagules and plants in the 100× concentrate treatmentreceived 2.04 g of MycoApply Ultra or 583.44 mycorrhizal propagules.

Percent mycorrhizal colonization produced significant differences(α=0.05) between treatment groups and the control (Table 2). Plantsgrown in the 100× concentrates treated with bactericide, both 7 days oldand 24 hours old, had a significantly higher percent mycorrhizalcolonization than plants grown in the 100× concentrate untreated, the10× concentrate untreated, and the control (Table 2). Plants grown inthe 7 days old 100× concentrate treated with bactericide had asignificantly higher percent mycorrhizal colonization compared to plantsgrown in the 10× concentrates treated with bactericides (Table 2).Plants grown in the 10× concentrates, both 7 days old and 24 hours old,had significantly higher percent mycorrhizal colonization compared tocontrol plants (Table 2).

For Trial 2 percent mycorrhizal colonization was not statisticallydifferent (α=0.05) between treatment groups and the control (Table 3).The plants grown in the 100× concentrate treated with 500 ppm Kathon and250 ppm Bronopol 24 hours old had the highest percent colonization andwas statistically different from all treatment groups except the 100×concentrate with the same bactericides at 7 days old (Table 3). All 100×concentrate treatment groups that contained bactericide treated MU hadstatistically higher percent colonization than the untreated 100×concentrate (Table 3). All 10× concentrate treatment groups thatcontained bactericide treated MU had statistically higher percentcolonization than the untreated 10× concentrate (Table 3). All treatmentgroups that contained bactericide treated MU had a statistically higherpercent colonization than the control (Table 3).

TABLE 2 Biocide Effect on Mycorrhizal Colonization Study I Mean %Standard Treatment Colonization Deviation Control 3.00 +/−3.71 c 10XMycoApply Ultra with 23.93 +/−9.12 b 500 ppm Kathon & 250 ppm Bronopolmade 7 days before treatment 10X MycoApply Ultra with 18.40 +/−15.81 b 500 ppm Kathon & 250 ppm Bronopol made 1 day before treatment 10XMycoApply Ultra with no biocides 9.93 +/−3.61 c 100X MycoApply Ultrawith 40.47 +/−8.17 a 500 ppm Kathon & 250 ppm Bronopol made 7 daysbefore treatment 100X MycoApply Ultra with 34.33 +/−6.51 a 500 ppmKathon & 250 ppm Bronopol made 1 day before treatment 100X MycoApplyUltra with no biocides 7.27 +/−3.76 c

TABLE 3 Biocide Effect on Mycorrhizal Colonization Study II Mean %Standard Treatment Colonization Deviation Control 5.07 +/−2.97 de 10XMycoApply Ultra with 19.93 +/−1.23 c 500 ppm Kathon & 250 ppm Bronopolmade 7 days before treatment 10X MycoApply Ultra with 17.73 +/−5.02 c500 ppm Kathon & 250 ppm Bronopol made 1 day before treatment 10XMycoApply Ultra with 21.00 +/−4.64 ac 500 ppm Kathon only made 1 daybefore treatment 10X MycoApply Ultra with no biocides 9.40 +/−2.77 de100X MycoApply Ultra with 28.40 +/−1.87 ac 500 ppm Kathon & 250 ppmBronopol made 7 days before treatment 100X MycoApply Ultra with 18.42+/−4.87 c 500 ppm Kathon & 250 ppm Bronopol made 1 day before treatment100X MycoApply Ultra with 34.53 +/−3.07 ab 500 ppm Kathon only made 1day before treatment 100X MycoApply Ultra with no biocides 14.67 +/−6.89cdConclusion:

In Trial 1 the treatment of MycoApply Ultra with 500 ppm Kathon and 250ppm Bronopol had a positive affect on the mycorrhizal infectivity ofMycoApply Ultra. The 100× MycoApply Ultra treated with bactericide had 5times greater mycorrhizal infectivity than 100× MycoApply Ultrauntreated. The 10× MycoApply Ultra treated with bactericide had 2 timesmore mycorrhizal infectivity compared to the 10× MycoApply Ultrauntreated.

In Trial 2 the treatment of MycoApply Ultra with 500 ppm Kathon and 250ppm Bronopol and 500 ppm Kathon had a positive effect on the mycorrhizalinfectivity of MycoApply Ultra. Trial 2 was set up to duplicate theresults obtained from Trial 1. The results of Trial 2 mimic the resultsobtained from Trial 1 and show that treatment with Kathon and Bronopolincrease mycorrhizal infectivity.

The results from both trials indicate that the presence of bactericide,specifically Kathon and Bronopol, increase the ability of mycorrhizas togerminate and infect roots.

The invention claimed is:
 1. A liquid composition, comprising (a) one ormore mycorrhizas; (b) an effective amount of a bactericide and/or abacteriostat for increasing mycorrhizal colonization; and (c) one ormore fertilizer ingredients.
 2. The liquid composition of claim 1,wherein the one or more mycorrhizas are ectomycorrhizas.
 3. The liquidcomposition of claim 1, wherein the one or more mycorrhizas areendomycorrhizas.
 4. The liquid composition of claim 1, wherein the oneor more mycorrhizas are ericoid mycorrhizas.
 5. The liquid compositionof claim 1, wherein the one or more mycorrhizas are arbutoidmycorrhizas.
 6. The liquid composition of claim 1, wherein the one ormore mycorrhizas are monotripoid mycorrhizas.
 7. The liquid compositionof claim 1, wherein the one or more mycorrhizas are orchid mycorrhizas.8. The liquid composition of claim 1, wherein the one or moremycorrhizas are a mixture of ectomycorrhizas and endomycorrhizas.
 9. Theliquid composition of claim 1, which comprises a bactericide.
 10. Theliquid composition of claim 9, wherein the bactericide is selected fromthe group consisting of a. 2-bromo-2-nitro-1,3-propanadiol, b.1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantane chloride, c.dibromonitrilopropionamide, d. 1,2-benzisothiazolin-3-one, e.5-chloro-2-methyl-4-isothiazolin-3-one,2-methyl-4-isothiazolin-3-one, f.diazolidinyl urea, g. tris(hydroxymethyl)nitromethane, h. sodiumo-phenylphenate, i. copper arsenates, j. cuprous oxide, k. compounds ofarsenic, copper, mercury, and l. quarternary ammonium compounds.
 11. Theliquid composition of claim 1, which comprises a bacteriostat.
 12. Theliquid composition of claim 11, wherein the bacteriostat is sodium azideor thimerosol.
 13. The liquid composition of claim 1, which comprises abactericide and a bacteriostat.
 14. The liquid composition of claim 1,wherein the one or more mycorrhizas are present in an amount of 0.2-5%.15. The liquid composition of claim 1, wherein the bactericide and/orthe bacteriostat is present in an amount of 50-3,000 ppm of the activeingredient.
 16. A method of increasing the colonization with one or moremycorrhizas, comprising applying a liquid composition of claim 1 to aplant, grass, tree or shrub.
 17. The liquid composition of claim 14,wherein the one or more mycorrhizas are present in an amount of0.33-3.3%.
 18. The liquid composition of claim 1, wherein thebactericide and/or the bacteriostat is present in an amount of 150-2,000ppm of the active ingredient.
 19. The liquid composition of claim 1,wherein the fertilizer ingredient is selected from the group consistingof a. Diammonium Phosphate, b. Potassium Nitrate, c. Urea, d. PotassiumPhosphate, e. Potassium Bicarbonate, f. Lidoquest Fe13P, g. LidoquestMn13P, h. LidoquestZn13P, i. Potassium Chloride (KCl), j. DisodiumDihydro Molybdate, k. Cobalt Chloride Hexahydrate, l. Nickel ChlorideHexahydrate, m. Caustic Potash Liquid, n. MKP, o. P-paraben/Nipasol, p.Potassium thiosulfate, and q. Sodium hydroxide.
 20. The liquidcomposition of claim 1, which further comprises a non-nutritional plantbeneficial substance or biostimulant, selected from the group consistingof a. Seaweed Extracts (Ascophyllum nodosum) b. Humic Acids, c. AscorbicAcid, d. Thiamine mononitrate, e. Potassium Sorbate (50% K Sorb), f.Myo-Inositol, g. Glycine, and h. Vitamin E (alpha tocopherol).